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Photoautotrophic Growth and Photosynthesis in Tobacco Callus Cells

Department of Biochemistry, The Connecticut Agricultural Experiment Station, New Haven, Connecticut 06504

Haploid tobacco (Nicotiana tabacum L.) cell cultures derived from quite different cultivars have been grown photoautotrophically in medium lacking sucrose and with 1.6 µM naphthaleneacetic acid and 1.5 µM isopentenylaminopurine. Cells were grown for 5 months on agar medium in Petri plates in air with dry weight increases of 1.5- to 3-fold per month. Callus cells were also grown photoautotrophically for at least three consecutive transfers 3 weeks apart in shallow liquid medium in horizontally placed gas-washing bottles where they were gassed continuously with air or air enriched with CO₂. Raising the CO₂ level in the air surrounding the cells increased the growth rate, and after about 3 weeks in 1% CO₂ the dry weight was approximately 3-fold greater than the inoculum. Growth rates remained about the same after each consecutive transfer. Autotrophic growth with this regime is not restricted to specific clones or cultivars.

Photosynthetic measurements in an atmosphere containing ¹⁴CO₂ established that rates of CO₂ assimilation in the callus cells at high CO₂ levels were similar to those of leaves on a chlorophyll basis, but were much slower on a fresh weight basis. Photosynthetic light saturation was achieved at an irradiation of about 125 µeinsteins m^–2 sec^–1 (400-700nm). The availability of photosynthetically dependent haploid cells provides an opportunity to select photosynthetic mutations which can be expressed in plants regenerated from these cells.

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